Copper imbalances, whether deficiency or overload, may result in the loss of cellular integrity and the disruption Of normal repair processes. Specific examples include the liver, where copper overload has been shown to result in cellular necrosis, and the pancreas, where copper deficiency can cause almost complete destruction of acinar cells and a striking loss of normal repair mechanisms. Thus, it is important that cellular copper concentrations be tightly regulated. Unfortunately, we do not have a clear understanding of the cellular and molecular mechanisms that are responsible for copper regulation. The focus of this project is the hypothesis that in response to fluctuations in dietary copper, there are changes in gene expression that are responsible for the maintenance of normal copper balance and cellular integrity. The experiments that are described here will include the identification and study of specific molecular mechanisms that play a role in the regulation of cellular copper levels. The Specific Aims will include isolation and study of novel copper-regulated genes as well as the examination of two recently identified genes, MNK and WD, apparently involved in copper regulation. Specific Aim I, Identification of Novel Copper Regulated Genes, will focus on the isolation, identification and cloning of novel copper regulated genes in the intestine, liver, brain and pancreas. Specific Aim II, Characterization of Novel Copper Regulated Clones, will examine the degree to which these novel genes are regulated by copper as well as other aspects of regulation such as developmental patterns of gene expression. Specific Aim III, Regulation of the MNK and WD) Genes, will examine the regulation of two-genes which encode putative copper transport proteins. Regulation of these genes during dietary copper restriction and overload will be explored with other aspects of regulation. Finally, Specific Aim IV, Role of MNK and WD) in Copper Transport, will use transient transfection of cultured cells to overexpress these genes and study their role in the tissue specific regulation of cellular copper levels. Together, these studies will provide information on the molecular regulation of copper and its role in the maintenance of cellular integrity and repair mechanisms.